The invention relates to methods of inhibiting the proliferation of cells and sensitizing cells to radiation therapy and DNA damaging chemotherapeutics, and in particular, treating cancer cells and individuals in vivo, including intra-operative treatments, by administration of Rad51 inhibitors including antisense molecules.
The control of the proliferation of cells is of interest. For example, inhibition of the proliferation of cells is useful in treating a number of disorders such as cancer, autoimmune disease, arthritis, inflammatory bowel disease, proliferation induced after medical procedures, and many other instances. Therefore, a number of approaches have been taken which are meant to inhibit the proliferation of cells. For example, chemotherapeutics are intended to inhibit proliferation or kill cancerous cells. However, while there have been many approaches to treating disorders requiring the inhibition of cell proliferation, there is still a need to identify more efficient treatments, particularly treatments which are sensitive and which have limited side effects.
In one approach, radiation is a major treatment mode for both children and adults with high grade gliomas. Although low linear energy transfer irradiation has been shown to have some beneficial effects on the treatment of astrocytic tumors, most malignant gliomas are radioresistant so that various methods of improving the therapeutic ratio in their treatment have been explored. The efficacy of fractionated irradiation, which is commonly employed in clinical practice, depends on four facts: redistribution of tumor cells in the cell cycle, repopulation, reoxygenation, and repair of sublethal damage. These factors have generated several approaches which have been applied in clinical practice. These include accelerated fractionation so as to reduce tumor repopulation, radiosensitization of hypoxic cells by hyperbaric oxygen and nitroimidazoles, and combination with chemotherapeutic agents such as BCNU and vincristine (9-11). So far, however, none of these procedures has resulted in satisfactory outcome for the treatment of malignant gliomas.
One study has reported that Rad51 antisense inhibition enhances radiosensitivity in normal cells, in vitro. Taki, et al., Biochemical and Biophysical Res. Comm., 223:434-438 (1996). However, this study does not report on the affects of Rad51 antisense inhibition in abnormal cells, such as tumor cells, nor does this study report on the affects of Rad51 antisense inhibition in vivo.
Rad51 is of interest because it is detected in every proliferating cell. It is believed that Rad51is within the family of proteins involved in repairing DNA damage, such as double-strand breaks in DNA caused by ionizing radiation and some alkylating agents, which lead to cell death if not repaired. Several genes related to double-strand break repair have been isolated from E. coli and S. cerevisia (Roca and Cox (1990); Shinohara (1992)). In most prokaryotes, including E. coli RecA protein or RecA-like protein plays an essential role in homologous recombination and in a variety of SOS responses to DNA damage (Kowalczykowski (1987)). In yeasts, which are lower eukaryotes, genes of the RAD52 epistasis group (RAD50-RAD57) have been identified by mutants not only as being deficient in their capability of DNA damage repair caused by ionizing radiation but also as having impaired capacity for mitotic and meiotic recombination (Resnick (1987); Friedberg (1988)). The Rad51 gene has been cloned and its product shown to be structurally similar to E. coli RecA protein with ATP-dependent DNA binding activity (Aboussekhara (1992); Basile (1992)). One study shows a mouse homologue of the yeast Rad51 gene that functionally complements a Rad51 mutation of S. cerevisiae with sensitivity to methylmethanesulfonate, a double-strand breaking agent (Morita (1993)).
The present invention, for the first time, provides methods to inhibit cell proliferation comprising administration of a Rad51 inhibitor. The invention further provides Rad51 inhibitor molecules that disrupt mammalian double stranded break repair. Moreover, the invention provides methods to treat diseased cells or individuals by administering a composition comprising a Rad51 inhibitor. Furthermore, the invention provides methods of inhibiting Rad51 expression in vivo using Rad51 inhibitors. Additionally, the invention provides methods of inducing sensitization to radiation, aklylating agents and other DNA damaging chemotherapeutics in vivo using Rad51 inhibitors. Also, the invention provides Rad51 inhibitors that are antisense molecules. Other aspects of the invention are described below.
The present invention provides methods for inhibiting cell proliferation in an individual comprising administering to the individual a composition comprising a Rad51 inhibitor. Also provided herein is a method for inhibiting the growth of a cell comprising administering to said cell a composition comprising a Rad51 inhibitor. Such methods can further include the step of providing radiation or alkylating agents after administration of said Rad51 inhibitor. In preferred embodiments the methods are performed in vivo and/or on cancerous cells and can be used with intra-operative treatments.
In a another aspect, the present invention provides methods for inhibiting cell proliferation in an individual in vivo comprising administering to the individual a composition comprising a Rad51 antisense molecule. Also provided herein is a method for inhibiting the growth of a cancerous cell comprising administering to said cell a composition comprising a Rad51 antisense molecule.
In another aspect, provided herein is a method for inducing sensitivity to radiation and DNA damaging chemotherapeutics in an individual in vivo comprising administering to said individual a composition comprising a Rad51 antisense molecule. Also provided herein is method for inducing sensitivity to radiation and alkylating chemotherapeutics in a cancerous cell comprising administering to said cell a composition comprising a Rad51 antisense molecule. In one embodiment, the methods provided herein also include the step of administering radiation or alkylating chemotherapeutic agents to a cell.
In one aspect, the method of sensitizing a cell to radiation or DNA damaging agents comprises administering to a cell at least one antisense molecule having a sequence selected from the group consisting of AS4, AS5, AS6, AS7, AS8 and AS9. Also provided herein is a method of prolonging survival in an individual with cancer comprising administering to said individual at least one antisense molecule having a sequence selected from the group consisting of AS4, AS5, AS6, AS7, AS8 and AS9.
In a further aspect of the invention, administration of the compositions herein comprises localized delivery of said Rad51 antisense molecule. Moreover, said methods provided herein may further comprise radiation treatment and/or chemotherapeutic treatment of said patient.
Further provided herein are kits for diagnosing and/or treating cancer comprising a Rad51 antisense molecule. In one aspect, the kit is for adjunctive therapy for cancer. In a preferred embodiment, the kit comprises at least one of packaging, instructions, suitable buffers, controls, and pharmaceutically acceptable carriers.